Cell Wall Extract Research Articles

PSVIII-2 Effects of post-ruminal yeast cell wall extract on dry matter intake, gastrointestinal fermentation and permeability, and concentrations of white blood cell types in beef heifers exposed to feed restriction

Abstract The objective of this study was to evaluate whether post-ruminal provision of a yeast cell wall extract (YCW) affected gastrointestinal fermentation characteristics and permeability, and white blood cell concentrations for heifers exposed to feed restriction (FR). Ruminally cannulated beef heifers (n = 8) were used in a 2 × 2 crossover design. Treatments (TRT) included an abomasal infusion of a placebo (PLC; maltodextrin) or YCW derived from Saccharomyces cerevisiae (2 g/dose) infused at 0600 and 1800 h. Heifers were subject to a 5-d baseline period followed by 5 d of FR where dry matter intake (DMI) was restricted to 30% of ad libitum intake. Data collection included daily DMI, and rumen, fecal and blood samples. Total gastrointestinal and post-ruminal permeability were measured using a ruminal dose of Cr-EDTA and an abomasal dose of Co-EDTA during the baseline and FR phases. Data were analyzed to determine the effects of YCW provision and phase, and the 2-way interaction. There were no TRT or TRT × phase interactions except for fecal propionate concentration. By design, DMI during FR was 30% (P < 0.01) of that observed during baseline (4.1 vs. 13.4 kg/d, respectively). The ruminal concentrations of acetate (45.4 vs. 69.2 mmol/L), propionate (14.5 vs. 26.7 mmol/L), and butyrate (9.9 vs. 16.5 mmol/L) were reduced (P < 0.01) by 34, 46, and 52%, respectively, with FR relative to baseline. Fecal propionate concentration (interaction, P = 0.04) numerically increased (P = 0.12) from baseline to FR for PLC (7.7 vs. 9.6 mmol/L, respectively), while YCW did not change among phases; however, no means differed. Exposure to FR reduced fecal butyrate concentration by 28% (P = 0.03). Gastrointestinal tract permeability (P < 0.01; 2.3 and 2.5% for total and post-ruminal, respectively) was greater during FR compared with baseline, with 2.0% of the Cr-EDTA and 1.6% of the Co-EDTA excreted in urine. The concentrations of neutrophils (P = 0.03) and eosinophils (P < 0.01) were greater during FR relative to baseline (2.28, 1.27 and 0.53, 0.30 ×106 cells/mL, respectively). Exposure to FR altered ruminal SCFA concentrations, increased blood concentrations of neutrophils and eosinophils, and increased gastrointestinal permeability suggesting an inflammatory response; however, post-ruminal provision of a YCW did not mitigate the response to a feed restriction challenge.

483 DMI, gastrointestinal tract fermentation and permeability, and white blood cell responses in beef heifers provided post-ruminal yeast cell wall extract following feed restriction

Abstract The objectives of this study were to 1) evaluate gastrointestinal fermentation characteristics and permeability, and white blood cell concentrations as beef heifers recovered from a feed restriction (FR) challenge, and 2) determine whether post-ruminal provision of a yeast cell wall extract (YCW) modulated responses. Ruminally cannulated beef heifers (n = 8) were used in a 2 × 2 crossover design. Treatments (TRT) included an abomasal infusion of a placebo (PLC; maltodextrin) or YCW derived from Saccharomyces cerevisiae (2 g/dose) infused at 0600 and 1800 h. Heifers were subject to 5-d of FR (offering 30% of ad libitum DMI) followed by five 5-d recovery phases (REC1, REC2, REC3, REC4, and REC5). Daily DMI, and rumen, fecal, and blood samples were collected on d 5 of FR, REC1, REC3, and REC5 with an additional collection on d 2 of REC1. Total gastrointestinal and post-ruminal permeability were measured using a ruminal dose of Cr-EDTA and an abomasal dose of Co-EDTA, respectively, during FR, REC1, REC3, and REC5 with concurrent urine collection. Data were analyzed to determine the effects of YCW provision, day, and the 2-way interaction. There were no TRT or TRT×day effects for any variables except DMI (interaction, P = 0.04), where DMI did not differ during the FR challenge; however, heifers provided YCW had a greater increase by REC1 compared with PLC heifers without any means differing. DMI progressively increased from FR, to REC1, REC2, and REC5. Ruminal acetate concentration increased (P < 0.01) from FR (45.4 mmol/L) to REC1 d5 (60.1 mmol/L) and increased further to REC3 (67.3 mmol/L) with no change thereafter (68.3 mmol/L in REC5). Ruminal propionate concentration increased (P < 0.01) from FR (14.5 mmol/L) to REC1 d2 (27.7 mmol/L) and was not different thereafter. Similarly, butyrate concentration increased (P < 0.01) from FR (7.9 mmol/L) to REC1 (d 5; 15.0 mmol/L) and was not different thereafter. Fecal acetate, propionate, and butyrate concentrations were not affected by day (P ≥ 0.12). Total gastrointestinal and post-ruminal permeability were greatest during FR (P < 0.01; 2.6% and 2.2% of the infused marker was excreted in urine, respectively) and decreased to 1.6% and 1.1%, respectively, in REC3 followed by an increase in REC5 for total (2.0% of infused) but not post-ruminal (1.0% of infused) permeability. The concentration of neutrophils (P < 0.01) was greatest on REC1 d2 (4.84 ×106 μmol/mL) and eosinophil concentration (P < 0.01) was greatest during FR (0.53 ×106 μmol/mL), least on d 2 of REC1 and increased on d 5 of REC1 but was not different thereafter. WBC (P < 0.01) gradually decreased (P = 0.02) from FR and REC1 d2 to REC3 and REC5. Post-ruminal provision of YCW may accelerate increases in DMI early in the recovery from FR, yet treatment did not modulate ruminal or fecal SCFA, blood concentrations, or gastrointestinal permeability.

Redundant role of PAD2 and PAD4 in the development of cardiovascular lesions in a mouse model of Kawasaki disease vasculitis.

Kawasaki disease (KD) is the leading cause of acquired heart disease in children. While circulating neutrophils are increased and activated during acute KD, it is unclear whether neutrophils and neutrophil extracellular traps (NETs) contribute to the pathogenesis of KD. Peptidylarginine deiminase 4 (PAD4), an enzyme involved in protein citrullination and essential for NETs formation, is implicated in the pathogenesis of various diseases. Here, we used the Lactobacillus casei cell wall extract (LCWE)-induced mouse model of KD vasculitis to determine the contribution of PAD4 in KD vasculitis. We found that the pan-PADs inhibitor, Cl-amidine, significantly reduced LCWE-induced cardiovascular lesions, but neutrophil-specific Padi4 KO mice did not impact the development of KD vasculitis. While in vitro treatment of macrophages, which highly express Padi4, with Cl-amidine inhibited IL-1β secretion, macrophage-specific Padi4 KO mice did not reduce the lesions. Padi4-/- mice also developed KD vasculitis, AFM30a, a PAD2 inhibitor, significantly reduced KD vasculitis in Padi4-/- mice, indicating a compensatory role of PAD2 in PAD4 deficiency. We also identified several citrullinated proteins in macrophages with constitutively active NLRP3 inflammasome that were inhibited by Cl-amidine treatment, suggesting that protein citrullination participates in NLRP3 inflammasome activation. These data indicate a dispensable role for PAD4-dependent NETs formation, and a redundant role of PAD2 and PAD4 in this murine KD vasculitis. The cardioprotective effects of Cl-amidine to reduce the severity of murine KD vasculitis are not limited to PAD4 inhibition and may include decreased citrullination in the inflammasome pathway.

Evaluation of dietary yeast derived mannan oligosaccharide for pacific whiteleg shrimp Penaeus vannamei: Effects on growth performance, immune response, hepatopancreas morphology and resilience to infection against Vibrio parahaemolyticus

With trends towards low fishmeal and higher plant ingredient-based diets for shrimp, there is consequently greater demand on feed additives to bolster the protein and nutrient profile quality for achieving more efficiency. Also, there is a need to enhance innate immunity and resilience to infection under intensive shrimp production scenarios. This investigation tested incremental levels 0, 0.2; 0.4; 0.6 and 0.8 % of a commercial yeast cell wall extract mainly comprising β-glucan and mannan oligosaccharides (MOS) (YM, YeaMOS, Angel Yeast, Yichang, Hubei, China) for post-larval shrimp Penaeus vannamei in isonitrogenous and iso-lipidic diets with proteinaceous plant ingredients amounting to 50 % contribution of the diet formulation. After 60 d, growth and feed utilization performance were not significantly affected in shrimp in terms of weight gain or FCR, but due to much higher survival, total biomass in groups of shrimps fed with dietary YM was higher compared to the control treatment. A sequential specific pathogen challenge test with Vibrio parahaemolyticus at final dose of 104 CFU mL−1 resulted in a marked improvement in survival against the control group with much enhancement at the inclusion level of 0.4 % YM (p<0.05). Before challenged, the total haemocyte count (THC) and the relative gene expression of Prophenoloxidase (ProPO) were higher in the group of shrimps fed with YM. Likewise, after infection, the amount of THC and ProPO remained better in the group of shrimps fed with YM. However, the 0.8 % YM treatment did not significantly differ from the control for phagocytic activity and index. The histomorphology condition of the hepatopancreas of shrimp before the infection for all treatments appears normal without significant numbers of hepatopancreatic tubules (T) and epithelial cell vacuoles (V). After pathogen challenge, all shrimp exhibited severe necrotic cell damage and massive sloughing of hepatopancreatic tubule epithelial cells into the lumen. Overall, giving YM as feed additives can improve the shrimp biomass and health, with the use of 0.4 % inclusion levels in practical diets for shrimp P. vannamei become the optimum dose to develop more efficient aquafeed production.

Abstract Tu035: Targeted Downregulation of MKI67 Mediated by miR-10b-5p Contributes to the Pathogenesis of Kawasaki Vasculitis

Kawasaki disease is a systemic disorder of enigmatic origin, postulated to stem from an aberrant immune response possibly triggered by infectious agents. This condition prominently risks developing vasculitis, particularly in the coronary arteries, which may lead to aneurysm formation, a grave complication. To shed light on the mechanisms underpinning Kawasaki vasculitis, a thorough investigation was conducted on patients presented at emergency departments. Whole RNA sequencing was utilized to analyze samples from individuals with general infectious diseases, acting as controls, and those diagnosed with Kawasaki disease, forming the experimental group. This approach identified a set of 13 microRNAs (miRNAs) that exhibited increased levels in patients with Kawasaki disease. Following initial findings, the Lactobacillus casei cell wall extract (LCWE)-induced vasculitis animal model, a recognized analog for human Kawasaki vasculitis, was employed to validate the relevance of these miRNAs. Among them, miR-10b-5p was pinpointed as a critical regulator; its inhibition via antagomirs markedly mitigated inflammation near the aortic valve in the LCWE-induced model. To uncover the direct downstream targets of miR-10b-5p relevant to Kawasaki vasculitis, single-cell RNA sequencing was performed. This revealed alterations in human coronary artery endothelial cells (HCAECs) upon miR-10b-5p treatment. Specifically, a subpopulation of HCAECs, initially characterized by proliferation, shifted towards a pro-inflammatory cluster upon miR-10b-5p exposure. This transformation was marked by a decrease in the expression of the proliferation marker, Ki-67 (MKI67), in the pro-inflammatory cluster. Further whole RNA sequencing on HCAECs treated with miR-10b-5p confirmed a significant downregulation of MKI67. Subsequent experiments with siRNA-mediated knockdown of MKI67 resulted in an increase in C-X-C Motif Chemokine Ligand 8 (CXCL8) expression, a pivotal cytokine in inflammation. These findings suggest a novel signaling pathway involving miR-10b-5p, MKI67, and CXCL8, potentially serving as a therapeutic target for Kawasaki vasculitis management in future interventions.

Long-term cardiovascular inflammation and fibrosis in a murine model of vasculitis induced by Lactobacillus casei cell wall extract.

Kawasaki disease (KD), an acute febrile illness and systemic vasculitis, is the leading cause of acquired heart disease in children in industrialized countries. KD leads to the development of coronary artery aneurysms (CAA) in affected children, which may persist for months and even years after the acute phase of the disease. There is an unmet need to characterize the immune and pathological mechanisms of the long-term complications of KD. We examined cardiovascular complications in the Lactobacillus casei cell wall extract (LCWE) mouse model of KD-like vasculitis over 4 months. The long-term immune, pathological, and functional changes occurring in cardiovascular lesions were characterized by histological examination, flow cytometric analysis, immunofluorescent staining of cardiovascular tissues, and transthoracic echocardiogram. CAA and abdominal aorta dilations were detected up to 16 weeks following LCWE injection and initiation of acute vasculitis. We observed alterations in the composition of circulating immune cell profiles, such as increased monocyte frequencies in the acute phase of the disease and higher counts of neutrophils. We determined a positive correlation between circulating neutrophil and inflammatory monocyte counts and the severity of cardiovascular lesions early after LCWE injection. LCWE-induced KD-like vasculitis was associated with myocarditis and myocardial dysfunction, characterized by diminished ejection fraction and left ventricular remodeling, which worsened over time. We observed extensive fibrosis within the inflamed cardiac tissue early in the disease and myocardial fibrosis in later stages. Our findings indicate that increased circulating neutrophil counts in the acute phase are a reliable predictor of cardiovascular inflammation severity in LCWE-injected mice. Furthermore, long-term cardiac complications stemming from inflammatory cell infiltrations in the aortic root and coronary arteries, myocardial dysfunction, and myocardial fibrosis persist over long periods and are still detected up to 16 weeks after LCWE injection.

Hydrogen Gas Inhalation Treatment for Coronary Artery Lesions in a Kawasaki Disease Mouse Model.

Kawasaki disease (KD) is a syndrome primarily affecting young children, typically under the age of five, and is characterized by the development of acute vasculitis. Through extensive research conducted on both murine and human subjects, it has been demonstrated that heightened levels of reactive oxygen species (ROS) play a pivotal role in the development of KD, especial coronary artery lesions (CALs). Hydrogen gas exhibits potent antioxidant properties that effectively regulate ROS production and the inflammatory response. We used Lactobacillus casei cell wall extract (LCWE)-induced vasculitis in mice as an animal model of KD and treated the mice with hydrogen gas inhalation. We observed significant dilatation and higher Z scores in the left coronary artery (LCA) in D21 and D28 in mice after LCWE treatment compared to the control group (p < 0.001) and a significant resolution of LCA diameters (p < 0.01) and Z scores (p < 0.01) after treatment with inhaled hydrogen gas. We further demonstrated that serum IL-6 expression was higher in mice after LCWE treatment (p < 0.01) and IL-6 significantly decreased after inhaled hydrogen gas therapy (p < 0.001). According to our literature review, this is the first report where hydrogen gas inhalation has been demonstrated to be effective for the treatment of coronary artery dilatation in a KD murine model.

Mitochondrial DNA release via the mitochondrial permeability transition pore activates the cGAS-STING pathway, exacerbating inflammation in acute Kawasaki disease

BackgroundKawasaki disease (KD) is an immune vasculitis of unknown origin, characterized by transient inflammation. The activation of the cGAS-STING pathway, triggered by mitochondrial DNA (mtDNA) release, has been implicated in the onset of KD. However, its specific role in the progression of inflammation during KD's acute phase remains unclear.MethodsWe measured mtDNA and 2’3’-cGAMP expression in KD patient serum using RT-qPCR and ELISA. A murine model of KD was induced by injecting Lactobacillus casei cell wall extract (LCWE), after which cGAS-STING pathway activation and inflammatory markers were assessed via immunohistochemistry, western blot, and RT-qPCR. Human umbilical vein endothelial cells (HUVECs) were treated with KD serum and modulators of the cGAS-STING pathway for comparative analysis. Mitochondrial function was evaluated using Mitosox staining, mPTP opening was quantified by fluorescence microscopy, and mitochondrial membrane potential (MMP) was determined with JC-1 staining.ResultsKD patient serum exhibited increased mtDNA and 2’3’-cGAMP expression, with elevated levels of pathway-related proteins and inflammatory markers observed in both in vivo and in vitro models. TEM confirmed mitochondrial damage, and further studies demonstrated that inhibition of mPTP opening reduced mtDNA release, abrogated cGAS-STING pathway activation, and mitigated inflammation.ConclusionThese findings indicate that mtDNA released through the mPTP is a critical activator of the cGAS-STING pathway, contributing significantly to KD-associated inflammation. Targeting mtDNA release or the cGAS-STING pathway may offer novel therapeutic approaches for KD management.

Abstract 1117: Il-1 Signaling On Vascular Smooth Muscle Cells Accelerates Atherosclerosis After Murine Kawasaki Disease Vasculitis

Objective: We have previously shown that Lactobacillus casei cell wall extract (LCWE)-induced Kawasaki disease (KD) vasculitis significantly accelerates atherosclerosis in hypercholesterolemic mice on high fat diet. IL-1 signaling is known to play a key role in both KD vasculitis and in the development of atherosclerosis. Here, we investigated the contribution of IL-1 signaling on vascular smooth muscle cells (VSMCs) in KD vasculitis-induced acceleration of atherosclerosis. Methods: Tamoxifen-inducible VSMC-specific IL-1 receptor ( Il1r1) knockout ( Myh11 Cre -ERT2 Il1r1 Δ/Δ ) mice and Il1r1 fl/fl littermate controls, all on ApoE -/- background, were injected with either PBS or LCWE. Following induction of KD vasculitis for 2 weeks, mice were fed a tamoxifen diet for 2 weeks to induce Il1r1 deletion on VSMCs, before being exposed to 8 weeks of Western diet to promote atherosclerosis. Results: KD vasculitis was associated with a significant acceleration of atherosclerosis, as expected. Myh11 Cre -ERT2 Il1r1 Δ/Δ mice had significantly diminished atherosclerotic plaque size, lipid composition, macrophage infiltration and necrotic core formation in aortic root as well as diminished lipid accumulation in aorta en face measurements compared with Il1r1 fl/fl control mice despite similar cholesterol levels. We also observed that Myh11 Cre -ERT2 Il1r1 Δ/Δ mice had significantly diminished endothelial adhesion molecules VCAM-1 and ICAM-1 expression in the lesion area and serum monocyte chemotaxis protein-1 (MCP-1) level compared with Il1r1 fl/fl control mice, consistent with the reduction of macrophage infiltration that we observed. Conclusions: Our results suggest an important pathophysiologic link between IL-1 signaling, specifically on VSMCs, and subsequent acceleration of atherosclerosis in hypercholesterolemic mice following KD vasculitis. Thus, further studies are warranted on the role of IL-1 signaling not only in acute KD, but also in the subsequent vascular remodeling and complications following acute KD vasculitis including atherosclerosis.

Targeting the SIRT1 pathway to modulate inflammation in a murine model of Kawasaki Disease vasculitis

Abstract Kawasaki Disease (KD) is an acute febrile systemic vasculitis and the leading cause of acquired heart disease among children. Overactivation of the NLRP3 inflammasome and IL-1b production are critical for the development of cardiovascular lesions during KD. Sirtuin 1 (SIRT1), a nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylase, is crucial for responding to cellular stress, and regulates NLRP3 activation as well as clearance of damaged mitochondria in cardiovascular diseases. However, its role in KD pathophysiology remains unknown. Here, we used the Lactobacillus casei cell wall extract (LCWE) murine model of KD to determine the role of SIRT1 on the development of cardiovascular lesions. We show that Sirt1 expression is downregulated in LCWE-injected mice with a significant reduction of nicotinamide, a product resulting from NAD+ degradation by SIRT1. In addition, supplementation of mice with NAD+ precursors significantly reduced the development of LCWE-induced cardiovascular lesions. Furthermore, KD development was less severe in SIRT1 overexpressing mice but was exacerbated with the specific deletion of Sirt1 in vascular smooth muscle cells. Treatment of LCWE-stimulated coronary smooth muscle cells with a SIRT1 activator favored mitophagy by inhibiting ribosomal S6 phosphorylation. These results indicate an impaired NAD+/SIRT1 axis during LCWE-induced KD vasculitis, which could potentially be targeted to reduce cardiovascular lesions in KD.

Abstract 1116: IL-33 Promotes Cardiovascular Lesion Development In A Murine Model Of Kawasaki Disease By Boosting IL-1β Production

Kawasaki Disease (KD), a pediatric acute febrile systemic vasculitis, is the leading cause of acquired heart diseases in children. Coronary artery aneurysms (CAAs) occur in up to 25% of untreated children, which is reduced to 5% with intravenous immunoglobulin (IVIG) treatment. However, up to 20% of KD patients are refractory to IVIG and at higher risk of developing CAAs. This highlights the need to characterize the immune mechanisms mediating KD to develop more efficient therapies. IL-1β plays a key role in KD pathogenesis. IL-33, a member of the IL-1 cytokine family, is released upon inflammation and tissue damage and exerts its effects by binding to its receptor ST2 ( Il1rl1 ). Circulating levels of IL-33 are elevated in KD patients during the acute phase of the disease. However, if and how IL-33 contributes to cardiovascular lesion development in KD remains unknown. Using the Lactobacillus casei cell wall extract (LCWE) murine model of KD vasculitis, we observed increased Il33 mRNA expression in hearts and abdominal aorta aneurysms of LCWE-injected mice. While ST2 was detected by immunofluorescence in the hearts of both control and LCWE-injected mice, its expression was increased in the abdominal aortas of LCWE-injected mice. In contrast, IL-33 expression was only increased in inflamed cardiovascular tissues of LCWE-injected mice. Single-cell RNA-sequencing and flow cytometric analysis of abdominal aortas, as well as spatial transcriptomics of heart tissues from LCWE-injected mice, indicated that Il33 transcripts were expressed primarily by stromal cells, such as fibroblasts, endothelial cells, and vascular smooth muscle cells. On the other hand, Il1rl1 transcripts were highly expressed by tissue infiltrating immune cells, such as macrophages, eosinophils, and neutrophils. Blocking IL-33, using either Il33 -/- mice or an anti-IL-33 antibody, significantly attenuated LCWE-induced KD vasculitis. In vitro, IL-33 treatment of LCWE-stimulated bone marrow-derived macrophages boosted their IL-1β production. Overall, our results indicate that IL-33, produced by stromal cells, may promote LCWE-induced KD vasculitis by increasing the release of IL-1β by tissue infiltrating immune cells, and this axis could therapeutically be targeted in KD.

Control of Peach Leaf Curl with Foliar Applications of Plant Immunity Inducers and Insights in Elicitation of Defense Responses against Taphrina deformans.

Taphrina deformans is the causal agent of leaf curl, a serious peach disease which causes significant losses in peach production worldwide. Nowadays, in order to control plant diseases, it is necessary to adopt novel and low-cost alternatives to conventional chemical fungicides. These promising strategies are targeted at eliciting host defense mechanisms via priming the host through the consecutive application of plant immunity inducers prior to pathogen challenge. In this study, we investigated whether chitosan or yeast cell wall extracts could provide enhanced tolerance against leaf curl in two-season field trials. Furthermore, we addressed the possible molecular mechanisms involved beyond the priming of immune responses by monitoring the induction of key defense-related genes. The efficacy of spraying treatments against peach leaf curl with both inducers was significantly higher compared to the untreated control, showing efficacy in reducing disease severity of up to 62.6% and 73.9% for chitosan and yeast cell wall extracts, respectively. The application of chitosan in combination with copper hydroxide was more efficient in reducing disease incidence and severity, showing efficacy values in the range of 79.5-93.18%. Peach plantlets were also spray-treated with immunity inducers three times prior to leaf inoculation with T. deformans blastospores in their yeast phase. The relative expression levels of nine key defense and priming genes, including those encoding members of pathogenesis-related (PR) proteins and hub genes associated with hormone biosynthesis, were monitored by RT-qPCR across three days after inoculation (dai). The results indicate that pre-treatments with these plant immunity inducers activated the induction of genes involved in salicylic acid (SA) and jasmonate (JA) defense signaling pathways that may offer systemic resistance, coupled with the upregulation of genes conferring direct antimicrobial effects. Our experiments suggest that these two plant immunity inducers could constitute useful components towards the effective control of T. deformans in peach crops.

Inhibition of IL-1 Ameliorates Cardiac Dysfunction and Arrhythmias in a Murine Model of Kawasaki Disease.

Kawasaki disease (KD) is an acute febrile illness and systemic vasculitis often associated with cardiac sequelae, including arrhythmias. Abundant evidence indicates a central role for IL (interleukin)-1 and TNFα (tumor necrosis factor-alpha) signaling in the formation of arterial lesions in KD. We aimed to investigate the mechanisms underlying the development of electrophysiological abnormalities in a murine model of KD vasculitis. Lactobacillus casei cell wall extract-induced KD vasculitis model was used to investigate the therapeutic efficacy of clinically relevant IL-1Ra (IL-1 receptor antagonist) and TNFα neutralization. Echocardiography, in vivo electrophysiology, whole-heart optical mapping, and imaging were performed. KD vasculitis was associated with impaired ejection fraction, increased ventricular tachycardia, prolonged repolarization, and slowed conduction velocity. Since our transcriptomic analysis of human patients showed elevated levels of both IL-1β and TNFα, we asked whether either cytokine was linked to the development of myocardial dysfunction. Remarkably, only inhibition of IL-1 signaling by IL-1Ra but not TNFα neutralization was able to prevent changes in ejection fraction and arrhythmias, whereas both IL-1Ra and TNFα neutralization significantly improved vasculitis and heart vessel inflammation. The treatment of L casei cell wall extract-injected mice with IL-1Ra also restored conduction velocity and improved the organization of Cx43 (connexin 43) at the intercalated disk. In contrast, in mice with gain of function of the IL-1 signaling pathway, L casei cell wall extract induced spontaneous ventricular tachycardia and premature deaths. Our results characterize the electrophysiological abnormalities associated with L casei cell wall extract-induced KD and show that IL-1Ra is more effective in preventing KD-induced myocardial dysfunction and arrhythmias than anti-TNFα therapy. These findings support the advancement of clinical trials using IL-1Ra in patients with KD.

Meta-Analysis of the Effects of Yeast Cell Wall Extract Supple-Mentation during Mycotoxin Challenges on the Performance of Laying Hens.

A random-effects meta-analysis was conducted to investigate the effect of mycotoxins (MT) without or with the inclusion of yeast cell wall extract (YCWE, Mycosorb®, Alltech, Inc., Nicholasville, KY, USA) on laying hen performance. A total of 25 trials were collected from a literature search, and data were extracted from 8 of these that met inclusion criteria, for a total of 12 treatments and 1774 birds. Laying hens fed MT had lower (p < 0.05) body weight (BW) by -50 g, egg production by -6.3 percentage points, and egg weight by -1.95 g than control fed hens (CTRL). Inclusion of YCWE during the mycotoxin challenges (YCWE + MT) resulted in numerically greater (p = 0.441) BW by 12.5 g, while egg production and egg weight were significantly (p < 0.0001) higher by 4.2 percentage points and 1.37 g, respectively. Furthermore, economic assessment calculations indicated that YCWE may not only support hen performance but also resulted in a positive return on investment. In conclusion, mycotoxins can play a role in negatively impacting laying hen performance and profitability. Inclusion of YCWE in feed with mycotoxin challenges provided benefits to egg production and egg weight and may support profitability. As such, the inclusion of YCWE could play an important role in minimizing mycotoxin effects and in turn aid farm efficiency and profitability.

Heterozygous mutations in factor H aggravate pathological damage in a stable IgA deposition model induced by Lactobacillus casei cell wall extract.

Activation of complement through the alternative pathway (AP) has a key role in the pathogenesis of IgA nephropathy (IgAN). We previously showed, by intraperitoneal injection of Lactobacillus casei cell wall extract (LCWE), C57BL/6 mice develop mild kidney damage in association with glomerular IgA deposition. To further address complement activity in causing glomerular histological alterations as suggested in the pathogenesis of IgAN, here we used mice with factor H mutation (FHW/R) to render AP overactivation in conjunction with LCWE injection to stimulate intestinal production of IgA. Dose response to LCWE were examined between two groups of FHW/R mice. Wild type (FHW/W) mice stimulated with LCWE were used as model control. The FHW/R mice primed with high dose LCWE showed elevated IgA and IgA-IgG complex levels in serum. In addition to 100% positive rate of IgA and C3, they display elevated biomarkers of kidney dysfunction, coincided with severe pathological lesions, resembling those of IgAN. As compared to wild type controls stimulated by the same high dose LCWE, these FHW/R mice exhibited stronger complement activation in the kidney and in circulation. The new mouse model shares many disease features with IgAN. The severity of glomerular lesions and the decline of kidney functions are further aggravated through complement overactivation. The model may be a useful tool for preclinical evaluation of treatment response to complement-inhibitors.

Involvement of IL-17 A/IL-17 Receptor A with Neutrophil Recruitment and the Severity of Coronary Arteritis in Kawasaki Disease

PurposeTo assess the role of the interleukin (IL)-17 A/IL-17 receptor A (IL-17RA) in Kawasaki disease (KD)-related coronary arteritis (CA).MethodsIn human study, the plasma levels of IL-17 A and coronary arteries were concurrently examined in acute KD patients. In vitro responses of human coronary endothelial cells to plasma stimulation were investigated with and without IL-17RA neutralization. A murine model of Lactobacillus casei cell-wall extract (LCWE)-induced CA using wild-type Balb/c and Il17ra-deficient mice were also inspected.ResultsThe plasma levels of IL-17 A were significantly higher in KD patients before intravenous immunoglobulin therapy, especially in those with coronary artery lesion. The pre-IVIG IL-17 A levels positively correlated with maximal z scores of coronary diameters and plasma-induced endothelial mRNA levels of chemokine (C-X-C motif) ligand-1, IL-8, and IL-17RA. IL-17RA blockade significantly reduced such endothelial upregulations of aforementioned three genes and inducible nitric oxide synthase, and neutrophil transmigration. IL-17RA expression was enhanced on peripheral blood mononuclear cells in pre-IVIG KD patients, and in the aortic rings and spleens of the LCWE-stimulated mice. LCWE-induced CA composed of dual-positive Ly6G- and IL-17 A-stained infiltrates. Il17ra-deficient mice showed reduced CA severity with the fewer number of neutrophils and lower early inducible nitric oxide synthase and chemokine (C-X-C motif) ligand-1 mRNA expressions than Il17ra+/+ littermates, and absent IL-17RA upregulation at aortic roots.ConclusionIL-17 A/IL-17RA axis may play a role in mediating aortic neutrophil chemoattraction, thus contributory to the severity of CA in both humans and mice. These findings may help to develop a new therapeutic strategy toward ameliorating KD-related CA.

The effects of ultrasonic processing on the antioxidant activity of Geotrichum candidum LG-8 and its cell wall extracts.

Extraction techniques that influence cell wall polysaccharides (EPS) is crucial for maximizing their bioactivity. This study evaluates ultrasound technology for extracting antioxidant polysaccharides from Geotrichum candidum LG-8, assessing its impacton antioxidant activity. Ultrasound extraction of EPS from G. candidum LG-8 was optimized (18 min, pH 7.0, 40 W/cm2, 0.75 M NaCl). ABTS scavenging efficiency and monosaccharide composition of LG-EPS1 and LG-EPS3 were analyzed using Fourier-transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). The Results showed that ultrasonic treatment markedly increased the ABTS radical scavenging efficiency of LG-8 cells by 47%. At a concentration of 1 mg/mL, the ultrasonically extracted LG-EPS1 and LG-EPS3 polysaccharides exhibited significant ABTS radical scavenging efficiencies of 26% and 51%, respectively. Monosaccharide composition analysis identified mannose and glucose in LG-EPS1, while LG-EPS3 was primarily composed of mannose. FTIR spectra verified the polysaccharides' presence, and SEM provided visual confirmation of the nanoparticle structures characteristic of LG-EPS1 and LG-EPS3. This research not only underscores the technological merits of ultrasound in polysaccharide extraction but also highlights the potential of G. candidum LG-8 derived polysaccharides as valuable bioactive compounds for antioxidant utilization.

Atomic Force Microscopy in the Characterization of the Structure of Cell Wall Components.

Atomic force microscopy (AFM) has broken boundaries in the characterization of the supramolecular architecture of cell wall assemblies and single cell wall polysaccharides at the nanoscale level. Moreover, AFM provides an opportunity to evaluate the mechanical properties of cell wall material which is not possible with any other method. However, in the case of plant tissue, the critical step is a smart sample preparation that should not affect the polysaccharide structure or assembly and on the other hand should consider device limitations, especially scanner ranges. In this chapter, the protocols from the sample preparation, including isolation of cell wall material and extraction of cell wall polysaccharide fractions, through AFM imaging of polysaccharide assemblies and single molecules until an image analysis to obtain quantitative data characterizing the biopolymers are presented.

LAMMER Kinase Governs the Expression and Cellular Localization of Gas2, a Key Regulator of Flocculation in Schizosaccharomyces pombe.

It was reported that LAMMER kinase in Schizosaccharomyces pombe plays an important role in cation-dependent and galactose-specific flocculation. Analogous to other flocculating yeasts, when cell wall extracts of the Δlkh1 strain were treated to the wild-type strain, it displayed flocculation. Gas2, a 1,3-β-glucanosyl transferase, was isolated from the EDTA-extracted cell-surface proteins in the Δlkh1 strain. While disruption of the gas2+ gene was not lethal and reduced the flocculation activity of the ∆lkh1 strain, the expression of a secreted form of Gas2, in which the GPI anchor addition sequences had been removed, conferred the ability to flocculate upon the WT strain. The Gas2-mediated flocculation was strongly inhibited by galactose but not by glucose. Immunostaining analysis showed that the cell surface localization of Gas2 was crucial for the flocculation of fission yeast. In addition, we identified the regulation of mbx2+ expression by Lkh1 using RT-qPCR. Taken together, we found that Lkh1 induces asexual flocculation by regulating not only the localization of Gas2 but also the transcription of gas2+ through Mbx2.

Role and mechanisms of CHI3L1 in coronary artery lesions in a mouse model of Kawasaki disease-like vasculitis

To explore the role and potential mechanisms of chitinase-3-like protein 1 (CHI3L1) in coronary artery lesions in a mouse model of Kawasaki disease (KD)-like vasculitis. Four-week-old male SPF-grade C57BL/6 mice were randomly divided into a control group and a model group, with 10 mice in each group. The model group mice were intraperitoneally injected with 0.5 mL of lactobacillus casei cell wall extract (LCWE) to establish a mouse model of KD-like vasculitis, while the control group mice were injected with an equal volume of normal saline. The general conditions of the mice were observed on the 3rd, 7th, and 14th day after injection. Changes in coronary artery tissue pathology were observed using hematoxylin-eosin staining. The level of CHI3L1 in mouse serum was measured by enzyme-linked immunosorbent assay. Immunofluorescence staining was used to detect the expression and localization of CHI3L1, von Willebrand factor (vWF), and α-smooth muscle actin (α-SMA) in coronary artery tissue. Western blot analysis was used to detect the expression of CHI3L1, vWF, vascular endothelial cadherin (VE cadherin), Caspase-3, B cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), nuclear factor κB (NF-κB), and phosphorylated NF-κB (p-NF-κB) in coronary artery tissue. The serum level of CHI3L1 in the model group was significantly higher than that in the control group (P<0.05). Compared to the control group, the expression of CHI3L1 in the coronary artery tissue was higher, while the expression of vWF was lower in the model group. The relative expression levels of CHI3L1, Bax, Caspase-3, NF-κB, and p-NF-κB were significantly higher in the model group than in the control group (P<0.05). The relative expression levels of vWF, VE cadherin, and Bcl-2 were lower in the model group than in the control group (P<0.05). In the LCWE-induced mouse model of KD-like vasculitis, the expression levels of CHI3L1 in serum and coronary arteries increase, and it may play a role in coronary artery lesions through endothelial cell apoptosis mediated by inflammatory reactions.